The mechanisms mediating kidney damage in diabetes mellitus undoubtedly stem from chronic hyperglycemia, but the intermediary steps are not completely understood. Evidence is accumulating that increased nonenzymatic glycation of proteins represents a mechanistic link between hyperglycemia and renal pathobiology. Glucose-derived modifications of proteins alter their functional and structural properties. In vivo, circulating glycated proteins principally exist as Amadori products (as opposed to advanced glycation products), and their concentration is significantly increased in diabetes with exposure to a hyperglycemic milieu. Recent focus on pathophysiologic events induced by advanced glycation has shifted attention from the possible role of Amadori- modified proteins in the development of diabetic complications. However, our published data with mesangial cells in culture and in the db/db diabetic mice have show that: Amadori-modified glucose adducts in albumin (GA) induce significant alterations in glomerular cell biology that resemble those of transforming growth factor-beta (TGF-b), a multi- functional cytokine with potent anti-proliferative and pro-fibrogenic activity; increased GA in diabetes is linked to increased bioactivity of the TGF-b/TGF-b receptor system; and neutralization of biologically active epitopes in GA ameliorates the structural and functional abnormalities characteristic of diabetic renal disease in db/db mice. The general plan of this project is to further investigate the role of GA and its molecular mediators in the increased accumulation of extracellular matrix (ECM) and the decrement in renal function characteristic of diabetic nephropathy. The Specific Aims are 1) to establish that increased expression of TGF-b1 is required for GA- stimulated ECM production; 2) to investigate that mechanism(s) underlying the increased expression of the TGF-b signaling receptors in mesangial cells grown with GA; 3) to examine the nature of the intracellular signal that mediates the stimulatory effect of GA on the production of TGF-b1; and 4) to establish that GA mediates up-regulation of the TGF-b1/TGF-b receptor system in the kidney and promotes long-term renal pathobiology in diabetic mice. A major component of these studies will be interventional arms in which we will administer neutralizing anti-GA and anti-TGF-b murine monoclonal antibodies to diabetic mice to prove that mesangial ECM expansion is caused by up-regulation of the renal TGF-b system. Understanding the mechanisms underlying up- regulation of the renal TGF-b system resulting from increased concentrations of GA may lead to the conceptual design of novel therapeutic interventions that could prevent the deleterious effects of diabetes on the kidney.